The immunotherapeutic approach to the treatment of cancer is based on the observation that human tumor cells express a variety of tumor-associated antigens (TAAs) that are not typically expressed in normal tissues. These antigens, which include viral tumor antigens, cellular oncogene proteins, and tumor-associated differentiation antigens, can serve as targets for the host immune system and elicit responses which result in tumor destruction. This immune response is mediated primarily by lymphocytes; T cells in general and class I MHC-restricted cytotoxic T lymphocytes in particular play a central role in tumor rejection. Hellstrom, K. E., et al., (1969) Adv. Cancer Res. 12:167–223; Greenberg, P. D. (1991) in Advances in Immunology, vol. 49 (Dixon, D. J., ed.), pp. 281–355, Academic Press, Inc., Orlando, Fla. Unfortunately, as evidenced by the high incidence of cancer in the population, the immune response to neoplastic cells often fails to eliminate tumors. The goal of active cancer immunotherapy is the augmentation of anti-tumor responses, particularly T cell responses, in order to effect complete tumor destruction.
Most attempts at active immunization against cancer antigens have involved whole tumor cells or tumor fragments. However, the cloning of TAAs recognized by CD8+ T cells has opened new possibilities for the immunotherapy of cancer based on the use of recombinant or synthetic anti-cancer vaccines. Boon, T., et al., (1994) Annu. Rev. Immunol. 12:337–365; Brithcard, V., et al., (1993) J. Exp. Med. 178:489–495; Cox, A. L., et al., (1994) Science 264:716–719; Houghton, A. N. (1994) J. Exp. Med. 180:1–4; Pardoll, D. M. (1994) Nature 369:357–358; Kawakami, Y., et al., (1994) Proc. Natl. Acad. Sci. U.S.A. 91:3515–3519; Kawakami, Y., et al., (1994) Proc. Natl. Acad. Sci. U.S.A. 91:6458–6462.
DF3/MUC1 (MUC1) is a cell surface glycoprotein that is overexpressed in breast, ovarian, and pancreatic tumors. The major extracellular portion of MUC1 is composed of tandem repeat units of 20 amino acids which comprise immunogenic epitopes. The full length major extracellular MUC1 protein is composed of up to 100 tandem repeat units of 20 amino acids containing 0-glycosylation sites which act as a framework for the formation of a highly glycosylated structure, which is highly immunogenic.
The term “tandem repeat unit” of MUC1 refers to the 20 amino acid repeated sequence of MUC1 (see, e.g., Gendler, S. J., et al (1990) J. Biol. Chem. 265:15286–15293).
(SEQ ID NO:1) GSTAPPAHGVTSAPDTRPAP
There is an abnormal glycosylation pattern found in carcinoma cells making the tumor-derived mucin antigenically distinct from normal mucin. Monoclonal antibodies specific for these peptide epitopes as well as their unique sugar side chains can identify >90% of breast tumors.
See Kufe, D., et al. (1984) Hybridoma 223–32; Taylor-Papadimitriou, J., et al. (1994) Trends Biotechnol. 12:227–33; Fontenot, J. D., et al. (1993) Cancer Res. 53:5386–94; Siddiqui J., et al. (1988) Proc. Natl. Acad. Sci. USA 85:2320–3; Merlo et al. (1989) Cancer Res. 49:6966–6971; and Abe, M., et al. (1989) Biochem Biophys Res Commun 165:644–9.
Accordingly, using the MUC1 tumor-associated antigen (TAA) has been proposed in developing cancer vaccines, particularly against tumors expressing MUC1. Multiple copies of tandem repeats are required for optimal native conformation and immunogenicity (see Fontenot et al., supra). A comparison of synthetic peptides containing 3, 4, or 5.25 tandem repeats of MUC1 revealed that the 5.25-copy version most closely mimicked the native structure of MUC1 and showed the most anti-mucin reactivity (Kotera et al. (1994) Cancer Res. 54:2856–2860). Previous recombinant vaccinia viruses containing the MUC1 gene with numerous tandem repeats were found to be unstable; homologous recombination resulted in deletion of most of the repeats, reducing the efficacy of the vaccine. See, e.g., Acres, R. B., et al. (1993) J. Immunother. 14:136–43; Bu, D., et al. (1993) J. Immunother. 14:127–35; Hareuveni, M., et al. (1990) Proc. Natl. Acad. Sci. USA. 87:9498–502; and Finn O. J. et al. infra.
The use of recombinant vaccinia viruses for anti-tumor immunotherapy has been discussed. (Hu, S. L., Hellstrom, I., and Hellstrom K. E. (1992) in Vaccines: New Approaches to Immunological Problems (R. W. Ellis, ed) pp. 327–343, Butterworth-Heinemann, Boston.) Anti-tumor responses have been elicited using recombinant pox viruses expressing TAAs such as carcinoembryonic antigen (CEA) and prostrate specific antigen (PSA). (Muraro, R., et al., (1985) Cancer Res. 4S:5769–5780); (Kantor, 3., et al. (1992) J. Natl. Cancer Inst. 84:1084–1091); (Robbins, P. F., et al. (1991) Cancer Res. 51:3657–3662) (Kantor, 3., et al. (1992) Cancer Res. 52:6917–6925.) No toxicity with these vectors was observed.
In general, viral vaccines are believed to mediate tumor rejection by activating class I MHC-restricted T-cells, particularly cytotoxic T lymphocytes (CTLs). T-cell activation is often potentiated by providing a suitable immunomodulator, for example a T-cell co-stimulatory factor such as those of the B7 gene family. See e.g., Greenberg, P. D. (1991) in Advances in Immunology, Vol. 49 (Dixon, D. J., ed.), pp. 281–355, Academic Press, Inc., Orlando, Fla.; Fox B. A. et al. (1990) J. Biol. Response Mod. 9:499–511.
It would be useful to have a recombinant pox virus encoding a MUC1 fragment containing a number of tandemly repeated sequences that will generate a cytotoxic T-cell response to MUC1, but which is stable, undergoing minimal excision as a result of homologous recombination in the gene encoding MUC1. It would also be useful to provide the recombinant pox virus in a vaccine format which is capable of potentiating T-cell activity against such tumors, particularly established or pre-existing tumors expressing the MUC1 TAA.